ABSTRACT Amyloids are a class of proteins that spontaneously self-assemble into cross ?-sheet aggregates which have been associated with amyloidosis and neurodegeneration. However, recently, amyloids that are non-toxic and rather play a functional role in a mammalian biosynthetic pathway have been discovered challenging our current views on the sole role of amyloids in mammals to be cytotoxic. Here we propose that amelogenin, the main protein of the developing enamel matrix, adapts cross-? sheet configuration and develops into fibrillar amyloids to achieve structural support to guide mineralization in vivo and in vitro. Enamel, the hardest and most mineralized tissue in the human body, is comprised of a unique organization of apatite nanofibers of about 50 nm in diameter and several hundreds of micrometers in length. There is agreement that the unique crystal morphology and organization into rod and interrod enamel is the result of a protein-guided uniaxial growth process of apatite, but it is unclear by which molecular mechanisms this unique micro- and nanostructure develops. While the role of self-assembly of enamel matrix proteins, in particular amelogenin, has widely been recognized as a crucial factor in controlling the structural development of enamel, a convincing relationship between organic supramolecular aggregates and enamel structure has only recently been observed, when we discovered that the recombinant human full-length amelogenin protein (rH174) forms ribbons of 17 nm in width, which grow to several micrometers in length. Such ribbons have the ability to self- align and form bundles resembling the appearance of aligned apatite crystallites in an enamel rod. Analysis of the primary structure of amelogenin revealed several domains with high propensity to form ?-sheets, including the possibilty to form amyloids, and produced a 14 residue N-terminal peptide (14P2) that readily assembled into nanoribbons (6.7nm wide), with possible amyloid structure. Amyloid stains were positive in enamel from mice lacking the enamel-specific enzyme kallikrein 4 (KLK4). Both enamel tissue and recombinant amelogenin nanoribbons showed x-ray diffraction spacings at 4.7 characteristic of ? sheets and amyloids. Enzymatic processing of self-assembled amelogenin promoted precise cleavage into the 23 kDa and 20 kDa fragments which resisted further degradation by MMP-20, thus possibly providing a stable organic template for mineralization during secretory stage amelogenesis, whereas the second enamel protease is able to disassemble and to degrade amelogenin amyloids. Herein we will further investigate the presence of amyloids in enamel and propose that amyloids play a key role in the development and organization of the organic matrix of enamel and that an exploration of such structures is essential to our understanding of enamel formation, its diseases and repair.